H. James Norton

Anti-Apoptotic Effects of IGF-1 and PDGF on Human Intervertebral Disc Cells In Vitro

Study Design. Human cells from the anulus were grown in tissue culture in an experimental design to study the anti-apoptotic effect of two selected cytokines. Objectives. To determine whether two selected cytokines, insulin-like growth factor-1 and platelet-derived growth factor, were effective in decreasing apoptosis in human cells from the anulus grown in culture for 10 days. Summary of the Background Data. Previous studies have shown that there is a small cell population in the aging human intervertebral disc. Earlier work from the authors’ laboratory suggested that apoptosis (programmed cell death) may be a major contributing factor to the decrease in cell number. A wide variety of inhibitors of apoptosis have now been identified; the present report presents findings on the actions of insulin-like growth factor-1 and platelet-derived growth factor in retarding or preventing apoptosis. Methods. Using previously published culture methods, cells from the anulus of 25 subjects (mean age, 41.7 years) were grown in monolayer culture for 10 days and tested under the following conditions: 1) control growth in the presence of 20% fetal bovine serum; 2) positive control conditions promoting the development of apoptosis in the absence of serum; or 3) in dose–response regimes where insulin-like growth factor-1 or platelet-derived growth factor were added in the presence of only 1% fetal bovine serum (necessary for basal cell maintenance). Specimens were derived from 18 lumbar, 9 cervical, and 1 thoracic sites; the average Thompson score was III. Cells were grown on chambered slides and evaluated in situ using the TdT in situ apoptosis detection reaction to identify apoptotic cells. An average of 300 cells were counted in replicate cultures at each dose to determine the incidence of apoptosis; results were analyzed with standard statistical techniques. Cultured cells also were examined with transmission electron microscopy. Results. Serum withdrawal to a 1% level was used as a positive apoptosis control in vitro and resulted in a significantly greater percentage of apoptosis compared with the 20% serum negative control (1.02% ± 0.34 (28) versus 0.14% ± 0.04 (27; mean ± SEM (n)), P < 0.0001). Exposure to 50 ng/mL insulin-like growth factor-1 significantly reduced the percentage of apoptosis (vs.— 1% serum) to 0.49% ± 0.26 (P = 0.005); 500 ng/mL was also significantly effective (% apoptosis = 0.09% ± 0.04 (P = 0.0001). Platelet-derived growth factor at a dose of 100 ng/mL also significantly reduced apoptosis (0.18 ± 0.11, P = 0.0001). Conclusions. Data demonstrate a significant reduction in the percentage of apoptotic disc cells after exposure to 50–500 ng/mL insulin-like growth factor-1 or exposure to 100 ng/mL platelet-derived growth factor. These findings expand the understanding of the cell biology of the disc cell and show that selected cytokines can retard or prevent programmed cell death in vitro. The administration of these cytokines may have future therapeutic potential in the treatment of disc degeneration.

Impact of Amoxicillin Prophylaxis on the Incidence, Nature, and Duration of Bacteremia in Children After Intubation and Dental Procedures

Background—Controversy exists about the impact of prophylactic antibiotics on bacteremia after invasive dental procedures. The purpose of this double-blind, randomized, placebo-controlled study was to determine the impact of amoxicillin prophylaxis on the incidence, nature, and duration of bacteremia from nasotracheal intubation and dental procedures in children. Methods and Results—Children were randomly assigned before surgery to the American Heart Association (AHA)–recommended dose of amoxicillin or to a placebo. Aerobic and anaerobic blood cultures were drawn at 8 specific time points after intubation, dental restorative and cleaning procedures, and before, during, and after dental extraction(s), to include blood drawings up to 45 minutes after the last extraction. Aerobic and anaerobic blood culture results were used to determine the incidence, nature, and duration of bacteremia from these procedures. For the 100 children enrolled (mean age, 3.5 years), the overall incidence of positive blood cultures, defined as at least 1 positive culture of the 8, was significantly higher in the placebo (84%) than the amoxicillin group (33%) (P <0.0001). Bacteremia occurrence rates after intubation and after dental restorations and cleaning were 18% and 20% in the placebo group and 4% and 6% in the amoxicillin group (P =0.05 and P =0.07, respectively). At 1.5 minutes after the initiation of dental extractions, bacteremia occurred in 76% of the placebo group versus 15% of the amoxicillin group (P <0.001). The majority of the 152 positive cultures and of the 29 different bacteria identified were Gram-positive cocci. Bacteremia persisted longer in the placebo group. Conclusions—Bacteremia from these procedures occurs more often, from a wider variety of bacterial species, and for a longer duration after dental extractions than previously reported in any age group. Amoxicillin has a significant impact on the incidence, nature, and duration of bacteremia after nasal intubation, dental restorative and cleaning procedures, and dental extractions.

Senescence in cells of the aging and degenerating intervertebral disc: Immunolocalization of senescence-associated β-galactosidase in Human and sand rat discs

Study Design. Human intervertebral disc anulus tissue was obtained in a prospective study of cell senescence. Localization of the senescence biomarker &bgr;-galactosidase (senescence associated &bgr;-galactosidase, SA-&bgr;-gal) was used for quantitative determination of the % senescent cells. Discs were obtained from surgical specimens or control donors. Discs were also studied from the lumbar spine of the sand rat. Experimental studies were approved by the authors’ Human Subjects Institutional Review Board and animal use committee. Objectives. To determine the incidence of cell senescence in human discs with Thompson Grades I through V using immunocytochemistry to quantify the percentage of cells positive for the senescence biomarker SA-&bgr;-gal. Summary of Background Data. Cell senescence has been recognized as a potential factor playing a role age-related disc degeneration. Senescent cells are viable but have lost the ability to divide. Senescence cells, however, are metabolically active. Methods. Fifty-seven discs specimens from 54 subjects were examined with immunocytochemistry for anti-SA-&bgr;-gal immunocytochemical localization to identify senescent cells. The fraction of positive cells was determined with quantitative histomorphometry. Results. Quantitative histomorphometry of human discs show an overall incidence of SA-&bgr;-gal-positive cells of 29.9% (±24.8, SD), with a range from 0 to 92.01%. Analysis by ANOVA of the % senescent cells grouped by Thompson grade showed significant increases in senescence with increasing disc degeneration (P < 0.0001). Further analysis with Tukey’s test showed significant differences between the % senescent cells in Grades I/II versus IV, and versus V. SA-&bgr;-gal-positive cells were also present in discs of the aging sand rat spine. Conclusions. Quantitative analysis of immunohistochemical localization of SA-&bgr;-gal identified a sizeable population of senescent cells in the aging/degenerating disc. It is important to discover more about the senescent disc cell population because these cells persist and accumulate over time within the disc. Since senescent cells cannot divide, senescence may reduce the disc’s ability to generate new cells to replace existing ones lost to necrosis or apoptosis.

Is surgical excision necessary for atypical ductal hyperplasia of the breast diagnosed by mammotome

BACKGROUND Core biopsy findings of atypical ductal hyperplasia (ADH) underestimates the diagnosis of malignancy by 18% to 88%. Using the Mammotome biopsy technique, more accurate assessment of the lesion is possible, making selective excision of these lesions a consideration. METHODS The records of 62 patients who were found to have ADH at Mammotome biopsy and subsequently underwent excision of the lesion were reviewed. Patient data were statistically analyzed for predictors of malignancy at the time of surgical excision. RESULTS Of the 62 patients, 9 (15%) had malignancy at excision. Variables predicting for malignancy included markedly atypical hyperplasia and incomplete removal of calcifications at Mammotome biopsy, a previous contralateral breast cancer, and a family history of breast cancer, with a combined sensitivity of 100% and specificity of 80%. CONCLUSIONS Mild ADH found on Mammotome, not associated with a personal or family history of breast cancer, may not need excision if all calcifications have been removed.

Decision-Making on the Use of Antimicrobial Prophylaxis for Dental Procedures: A Survey of Infectious Disease Consultants and Review

There is debate concerning use of antibiotic prophylaxis before invasive dental procedures for patients at risk of acquiring distant site infection (DSI). We determined the opinions and practices of infectious disease consultants (IDCs) regarding antimicrobial prophylaxis to prevent DSIs that result from invasive dental procedures by conducting a survey of the 797 members of the Infectious Diseases Society of America Emerging Infections Network (477 members [60%] responded). Ninety percent of respondents closely follow the American Heart Association guidelines for antibiotic prophylaxis for patients with valvular heart disease who undergo invasive dental procedures. In contrast, few IDCs recommend prophylaxis for patients with lupus erythematosus, poorly controlled diabetes mellitus, dialysis catheters or shunts, cardiac pacemakers, or ventriculoperitoneal shunts. Twenty-five percent to forty percent of respondents recommended prophylaxis for prosthetic vascular grafts, orthopedic implants, or chemotherapy-induced neutropenia. We conclude that IDCs differ considerably in their assessment of the need for prophylaxis for patients who have noncardiac risk factors for DSI. These differences underscore the need for definitive studies to delineate appropriate candidates for antimicrobial prophylaxis in dental practice.

Surrogate markers for adverse outcomes in normotensive patients with pulmonary embolism

Background:Although echocardiography has proven utility in risk stratifying normotensive patients with pulmonary embolism, echocardiography is not always available. Objective:Test if a novel panel consisting of pulse oximetry, 12-lead electrocardiography, and serum troponin T would have prognostic equivalence to echocardiography and to examine the prognostic performance of age, previous cardiopulmonary disease, D-dimer, brain natriuretic peptide, and percentage of pulmonary vascular occlusion on chest computed tomography. Design:Prospective cohort study. Patients and Setting:Normotensive (systolic blood pressure of >100 mm Hg) emergency department and hospital inpatients with diagnosed pulmonary embolism who underwent cardiologist-interpreted echocardiography and other measurements within 15 hrs of anticoagulation. Measurements and Main Results:End points were in-hospital circulatory shock or intubation, or death, recurrent pulmonary embolism, or severe cardiopulmonary disability (defined as echocardiographic evidence of severe right ventricular dysfunction with New York Heart Association class III dyspnea or 6-min walk test of <330 m) at 6-month follow-up. The two-one–sided test tested the hypothesis of equivalence with one-tailed &agr; = 0.05 and &Dgr; = 5%. Of 200 patients enrolled, data were complete for 181 (88%); 51 of 181 patients (28%) had an adverse outcome, including in-hospital complication (n = 18), death (n = 11), recurrent pulmonary embolism (n = 2), or cardiopulmonary disability (n = 20). Right ventricular dysfunction on initial echocardiogram was 61% sensitive (95% confidence interval, 46–74%) and 57% specific (48–66%). The panel was 71% sensitive (56–83%) and 62% specific (53–71%). The two-one–sided procedure demonstrated superiority of the panel to echocardiography for both sensitivity and noninferiority for specificity. No other biomarker demonstrated equivalence, noninferiority, or superiority for sensitivity and specificity. Conclusion:Normotensive patients with pulmonary embolism have a high rate of severe adverse outcomes during 6-month follow-up. A panel of three widely available tests can be used to risk stratify patients with pulmonary embolism when formal echocardiography is not available.

New Tumor-Node-Metastasis Staging Strategy for Node-Positive (stage III) Rectal Cancer: An Analysis

PURPOSE The tumor-node-metastasis system for staging rectal cancer is based on invasion, number of involved nodes, and metastasis. Nodes are classified as N1 or N2 according to the number involved with metastases. Nodal positivity defines stage III regardless of depth of invasion or number of positive nodes. Our purpose was to analyze overall survival when node-positive patients were stratified into three new subsets. METHODS We analyzed data entered into the National Cancer Data Base for 5,987 stage III patients with rectal cancer between 1991 and 1993. Survival was calculated using three new subgroups (IIIA: T1/2, N1; IIIB: T3/4, N1; IIIC: any T, N2). Survival following surgery and adjuvant therapy was assessed. The observed survival rates were calculated and compared using the log-rank method. The Cox regression model assessed subgroup differences. RESULTS Five-year observed survival rates for stage III subcategories were 55.1% in IIIA; 35.3% in IIIB; and 24.5% in IIIC. Stratifying for treatment outcome, stage IIIA patients having surgery alone (n = 278) had poorer observed 5-year survival (39%) than patients treated with surgery and adjuvant chemotherapy or radiation therapy (chemo/XRT; n = 765; 60%). Similar outcomes occurred in IIIB (surgery-alone [n = 726; 21.7%] and chemo/XRT [n = 2,130; 40.9%] groups) and in IIIC (surgery-alone [n = 467; 12.2%] and chemo/XRT [n = 1,621; 28.9%] groups). Differences were significant (P <.0001) in all stages. CONCLUSION The traditional stage III designation of rectal cancer fails to account for invasion (T1-4) and number of involved nodes (N1, N2). The stratification of stage III patients into three subsets should be used in future analyses of rectal cancer. The effect of postoperative adjuvant therapy was beneficial in all subsets.

The sand rat model for disc degeneration: radiologic characterization of age-related changes: cross-sectional and prospective analyses.

Study Design. This report is composed of two studies, one cross-sectional and one prospective cohort study, that analyze the radiologic features of disc degeneration in the sand rat (Psammomys obesus). Objectives. To statistically assess progressive disc degeneration in this useful animal model in terms of a cross-sectional study and a prospective monthly evaluation of individual animals. Summary of Background Data. P. obesus is an attractive small rodent model for spontaneous age-related disc degeneration. Because disc degeneration is spontaneous, the model avoids use of chemonucleolysis or surgical injury to cause disc degeneration. Little is understood, however, about specific details of the progressive disc deterioration. Methods. This study statistically assessed 158 animals in a cross-sectional study and 22 animals in a longitudinal study, which followed individual animals to 12 months of age. Radiologic features involving irregular disc margins, disc wedging, disc narrowing, endplate calcification, subchondral sclerosis, ligament calcification, and osteophyte formation were studied. Results. Significant age-related cross-sectional changes were present for all features (P ≤ 0.005). Males showed a statistically greater incidence of wedging at 6 and 12 months, wedging at 2 and 6 months, and endplate calcification at 2 months than did females. By 6 months of age, however, endplate calcification had a higher incidence in females than in males. Prospective analysis showed that wedging, narrowing, endplate calcification, and irregular disc margins were more common at 12 months of age than at 2 months (P = 0.0001). By the age of 12 months, all lumbar sites of both males and females showed endplate calcification and the majority of animals showed narrowing and wedging. Conclusions. Radiographic signs of degeneration were evident by age 2 months; wedging, narrowing, irregular disc margins, and endplate calcification were the most common degenerative changes in older animals. These data show that the sand rat provides a reliable, useful model of spontaneous disc degeneration.

Vertebral endplate architecture and vascularization: application of micro-computerized tomography, a vascular tracer, and immunocytochemistry in analyses of disc degeneration in the aging sand rat.

Study Design. Lower lumbar vertebral endplates from young and old sand rats were assessed in an Institutional Animal Care and Use Committee approved study for architectural endplate features using micro-computerized tomography (CT) 3-dimensional (3D) models and vascularization studies by an in vivo vascular tracer or immunocytochemical identification of blood vessels. Objective. To assess endplate porosity and vascularization using &mgr;CT architectural analysis, an in vivo vascular tracer, and immunocytochemical identification of blood vessels in the endplate. Summary of the Background Data. The vertebral endplates, also called cartilage endplates, form the superior and inferior, or cranial and caudal, boundaries of the disc. In the human being and sand rat, the cartilaginous endplate undergoes calcification with aging and is replaced by bone. Endplate sclerosis has long been thought to play a role in disc degeneration by decreasing nutrient availability to the disc, but this is still poorly understood. Previous work has identified increasing bone mineral density with aging and disc degeneration in the sand rat model. Methods. &mgr;CT models of the lower lumbar endplates of vertebrae at L5–6 and L6–7 were constructed from 6 younger (mean age 11 months) and 21 older (mean age 25.6 months) sand rats. Architectural features were scored on a semiquantitative scale for smoothness of the endplate face, irregularities on the endplate margin, and endplate thickness. There were 2 smaller sets of animals (n = 18) evaluated for endplate vascularity following in vivo injection of a fluorescent vascular tracer or by the use of immunocytochemistry to identify blood vessels. Results. &mgr;CT revealed a solid bony surface to the endplate, which was not penetrated by vasculature; with aging/disc degeneration, there was roughening and pitting of the plate surface, and the development of irregularmargins. In L5–6 and L6–7, sites of prominent disc degeneration evident on radiographs, the proportion of abnormalities in surface smoothness, margin irregularity, and endplate thickening were all statistically significant in both younger and older animals (P ≤ 0.0027). More severe changes were evident in the caudal versus cranial endplate surfaces. Histologic study of vascular tracer showed that there was no penetration of the disc by vascular supply from the endplate; this was verified by immunocytochemical identification of blood vessels. The canal system within the endplate was a complex 3D interconnected network. Conclusions. Findings show that disc degeneration in the sand rat occurs concomitantly with marked architectural bony changes on the endplate face, including loss of smoothness and development of irregular bony margins. Vascular connections were not present between the endplate and disc; this was verified with &mgr;CT studies, in vivo vascular tracers, and traditional immunocytochemistry. The canal system within the imaged endplate was revealed to consist of a complex 3D interconnected network.

EMT Defibrillation Does Not Increase Survival From Sudden Cardiac Death in a Two-Tiered Urban-Suburban EMS System☆☆☆★★★♢

OBJECTIVE The use of automatic external defibrillators (AEDs) by EMS initial responders is widely advocated. Evidence supporting the use of AEDs is based largely on the experience of one metropolitan area, with effect on survival in many systems not yet proved. We conducted this study to determine whether the addition of AEDs to an EMS system with a response time of 4 minutes for first-responder emergency medical technicians (FREMTs) and 10 minutes for paramedics would affect survival from cardiac arrest. METHODS This prospective, controlled, crossover study (AED versus no AED) of consecutive cardiac arrests managed by 24 FREMT fire companies took place from 1992 to 1995 in Charlotte, North Carolina, a city of 455,000. Patients were stratified using the Utstein criteria. The primary endpoint was survival to hospital discharge among patients with bystander-witnessed arrests of cardiac origin. RESULTS Of the 627 patients, 243 were bystander-witnessed arrests of cardiac origin. Survival to hospital discharge was accomplished in 5 of 110 patients (4.6%; 95% confidence interval [CI] 0.6% to 8.4%) with AED compared with 7 of 133 (5.3%, 95% CI 1.5% to 9.1%) without AED (P = .8). Both groups were comparable with regard to age, gender, history of myocardial infarction, congestive heart failure or diabetes, arrest at home, bystander CPR, and whether or not ventricular fibrillation (VF) was the initial rhythm. For arrests of any cause, witnessed by bystanders or EMS personnel, with an initial rhythm of VF or ventricular tachycardia (VT), 5 of 77 (6.5%, 95% CI 1.0% to 12.0%) with AED survived compared with 8 of 105 patients (7.6%, 95% CI 2.5% to 12.7%) without AED (P = .8). Statistically significant differences were noted in race and EMS response times between the two groups, which did not affect survival. CONCLUSION Addition of AEDs to this EMS system did not improve survival from sudden cardiac death. The data do not support routinely equipping initial responders with AEDs as an isolated enhancement, and raise further doubt about such expenditures in similar EMS systems without first optimizing bystander CPR and EMS dispatching.